Mass spectrometry-based spectral count has been a common choice of label-free proteome quantification due to the simplicity for the sample preparation and data generation. The discriminatory nature of spectral count in the MS data-dependent acquisition, however, inherently introduces the spectral count variation for low-abundance proteins in multiplicative LC-MS/MS analysis, which hampers sensitive proteome quantification. As many low-abundance proteins play important roles in cellular processes, deducing low-abundance proteins in a quantitatively reliable manner greatly expands the depth of biological insights. Here, we implemented the Moment Adjusted Imputation error model in the spectral count refinement as a post PLGEM-STN for improving sensitivity for quantitation of low-abundance proteins by reducing spectral count variability. The statistical framework, automated spectral count refinement by integrating the two statistical tools, was tested with LC-MS/MS datasets of MDA-MB468 breast cancer cells grown under normal and glucose deprivation conditions. We identified about 30% more quantifiable proteins that were found to be low-abundance proteins, which were initially filtered out by the PLGEM-STN analysis. This newly developed statistical framework provides a reliable abundance measurement of low-abundance proteins in the spectral count-based label-free proteome quantification and enabled us to detect low-abundance proteins that could be functionally important in cellular processes.
Bibliographical noteFunding Information:
This work was supported by the National Research Foundation of Korea (NRF) Grant funded by the Korean Government (MSIP) (NRF-2016R1A5A1010764 and NRF-2015M3A9B6073835 to ECY; NRF-2016R1D1A1B04931656 and NRF-2011-0025320 to KMK) and Global Infrastructure Program through the NRF funded by the Ministry of Science and ICT (NRF-2017K1A3A1A19071651 to ECY).
© 2019, The Author(s).
All Science Journal Classification (ASJC) codes